Diabetes Mellitus:- Part 5 – Glucose in Urine (Glycosuria), Urine Sugar, Benedict’s solution
- The test sample is urine.
- The best sample after 2 to 3 hours of the meal.
- To diagnose diabetes mellitus.
- To monitor the diabetes mellitus.
- To evaluate the effectiveness of the therapy.
- To diagnose gestational diabetes.
- It is part of a routine urine examination.
- Can see false-positive tests when any substance present in the urine can reduce the copper in the clinitest strips.
- Can see the false positive tests in the presence of other sugars like galactose, fructose, and lactose.
- Drugs giving false positive tests are acetylsalicylic acid, Ascorbic acid, cephalothin, chloral hydrate, streptomycin, sulphonamides, and aminosalicylic acid.
- Contamination from the oxidizing agents or bleach.
- Improper storage of the urine strips.
- Drugs giving false-negative results are levodopa, phenazopyridine, and ascorbic acid with clinitest strips.
- Multistix detects ≥50 mg/dL, if the amount is less then it will be negative.
- Sodium fluoride cause enzyme inhibition.
- Refrigerated specimens give decreased enzyme activity.
- There are drugs that give increased glucose in the urine like diuretics (thiazide), estrogens, isoniazid, lithium, nalidixic acid, nicotinic acid, chloramphenicol, chloral hydrate, cephalosporin, and aminosalicylic acid.
- Examination of urine for glucose is rapid, noninvasive, and inexpensive for screening the urine.
- A large number of urine samples can be tested.
- Glucose is filtered by the glomeruli and reabsorbed by proximal convoluted tubules.
- When glucose exceeds the renal threshold level(>180 mg/dL) of tubules then, glucose appears in the urine is called glycosuria.
- Tubular absorption is an active process to maintain the body’s glucose level.
- Renal threshold for glucose = 160 to 180 mg/dL.
- After the renal threshold values glucose appears in the urine.
- This is not sensitive nor specific for the control of diabetes because we don’t know about the glucose level below 180 mg/dL.
List of reducing substances in the urine are:
- Other substances are:
- Uric acid.
- Ketone bodies.
- Oxalic acid.
- Glucuronic acid.
- Hippuric acid.
- Homogentisic acid.
- drugs are:
- Ascorbic acid.
- Normally sugar (glucose ) is absent in the urine.
- Random specimen = negative
- 24 hours specimen = < 0.5 g/day (<2.78 mmol/day).
- Glucose appears in the urine when a blood glucose level of 180 mg/dL or more (cross the renal threshold).
- Its concentration in the urine correlates with the blood glucose level.
Procedure for glucose in the urine:
Benedict’s solution reagents:
- It consists of:
- CuSO4 (cupric sulphate) = 17.3 grams
- Sodium citrate (Na3C6H5O7-2H2O) = 173 grams
- Sodium carbonate (Na2CO3) = 100 grams
- Distle water = 1000 mL
- Procedure to prepare the solution:
- Dissolve cupric sulfate in hot water of 100 mL.
- Now dissolve separately sodium citrate and sodium carbonate with heating water of 800 mL. Let it cool.
- Mix solutions 1 and 2 and makeup to 1000 ml of water.
- Benedict’s reagent is ready and is stable.
- Benedict’s qualitative method is very common.
- It contains cupric ions complexed to citrate in an alkaline medium.
- Reducing substances convert cupric to cuprous ion.
- It forms yellow cuprous hydroxide or red cuprous oxide.
- Benedict’s solution method:
- This can be done on the solution of benedicts as well.
- Take 2.5 mL Benedict’s solution.
- Add 0.2 ml of the urine.
- Place the tubes in heat-block or heat directly to bring to 100 °C.
- Examine each tube color and the precipitate.
- Different colors develop according to the quantity of sugar in the urine.
- Greenish brown when there is a large quantity >2 g/dL.
- Another way to do Benedict’s reaction:
- Benedict’s solution 5 mL.
- Urine 0.4 mL (8 drops) .
- Mix and keep in a boiling water bath for 3 minutes.
- Tablet method:
- 5 drops of the urine are mixed with 10 drops of water.
- Then add the tablet as known clinitest.
- Procedure for Benedict reaction with tablet (Clinitest) shown in the following diagram.
- Also, run the negative control as well.
Reporting the result:
- It can be reported as plus + signs, from 1+ to 4+.
- It can report the percentage of 1 to 2 %. This reporting is more accurate.
|Color of the urine after Benedict’s reaction||Reporting method
||The concentration of glucose
|Blue, clear or cloudy (Benedict’s solution color)||0||NIL|
|Green and no precipitate (may see precipitate)||1+||Traces|
|Brown and cloudy||2+||Around 0.5 g%|
|Orange and cloudy||3+||Around 1.o g%|
|Red and cloudy||4+||Around 2.0 g% or more|
- A semiquantitative method where there are different strips is available like Clinistix, Diastix, and Chemstrip.
- In all the above strips, glucose-specific enzyme glucose oxidase is used.
- This is more specific for glucose than Benedict’s method.
- This test is positive when glucose concentration is 100 mg/dL or more.
- The sensitivity of the commercially available Discs:
- Multistix = 75 to 125 mg/dL
- Diastix = 75 to 125 mg/dL
- Chemstrip = 40 mg/dL in 90% of the specimen.
- The drawbacks of the strips:
- Urine strips detect mainly glucose, so there are chances for false-negative results due to interfering chemicals in the urine.
- False-positive results are seen in:
- If the container is contaminated by the detergents.
- In very dilute urine, traces may be seen due to sensitivity at low specific gravity.
- If the strips are exposed to air due to improper storage.
- False-negative results are seen in:
- It is seen in the intake of vitamin C (ascorbic acid), ad tetracyclines.
- In case if there are high ketone bodies (≥40 mg/dL) and low glucose level (75 to 125 mg/dL).
- Sodium fluoride inhibits the enzyme reaction, so it should not be used in the urine as a preservative.
- If urine is refrigerated then it will give false-negative results because of decreased enzyme activity, so bring the urine at room temperature
- The quantitative method uses hexokinase or glucose dehydrogenase procedure.
Increased glucose in urine seen in:
- Diabetes mellitus.
- Renal glycosuria.
- Hereditary defects in the metabolism of other reducing substances like galactose, pentose, and fructose.
- Liver diseases.
- Pancreatic diseases.
- Cushing’s syndrome.
- Brain injuries.
- Fanconi’s syndrome (Tubular defect).
- Advanced renal tubular diseases.
- Nephrotoxic chemicals like carbon monoxide, lead, and mercury.
The false-negative result is seen in:
- Mostly seen due to drugs.
- Ascorbic acid.
Comparison of Benedict reaction and Oxidase method:
Characteristics Benedict reaction (CuSO4) Glucose oxidase Minimum level detected Glucose 50 to 250 mg/dL Glucose 50 mg/dL Other sugars detected
Specific for glucose False-negative
- Ascorbic acid (Vitamin C)
- A large number of salicylates
- Homogentisic acid
- Vitamin C (Ascorbic acid)
- Many antibiotics
- X-ray contrast media
- Homogentisic acid
- Please see more details on Fasting Blood Glucose